Neural crest cells differentiate into sympathetic neurons and other cell types in response to specific tissue interactions or their environmental stimuli. Preliminary evidence suggests that these interactions might be mediated by one or more extracellular matrix (ECM) materials. Neural crest cells contribute extensively to connective tissues, glandular and neuronal elements of the head. Therefore, a study of ECM influence on neural crest cell differentiation is useful to the understanding of craniofacial development in addition to neuronal development. Chick neural crest cells require a specific sequence of interactions with ventral neural tube and somite tissue in order to differentiate in organ culture into sympathetic neurons, detectable by a specific formaldehyde-induced fluorescence assay for catecholamines. The proposed research is designed to further characterize these interactions: 1) fluorescent cells, which appear in organ culture, will be classified, by histological and diagnostic procedures, as prinicpal neurons or small intensely fluorescent (SIF) cells. 2) The timing of the competence of the various tissues in the organ culture system to support sympathetic neuron differentiation will be determined. 3) The relationship between ECM and sympathetic neuron differentiation in the organ culture system will be examined by using enzymes and other chemicals to interrupt or promote cartilage matrix formation. 4) A neural crest culture system will be developed which supports sympathetic neuron differentiation. This system will involve the deposition of ECM in a Millipore filter by cultured neural tube plus somite. 5) The ECM residues in the treated filters will be analyzed for glycosaminoglycans and collagen. 6) Various ECM's, hormones, growth factors, and neural crest cell interactions will be examined for effects on sympathetic neuron differentiation. 7) Cell culture conditions will be developed which will support growth of neural crest cells and then their differentiation into sympathetic neurons only in response to a regulated environmental stimulus.